The effect of
fungus on the diet
of broilers in Ahwaz
Abdoallah Beatsayah
, Seyed Asghar Khazaee
, Mohammad Chenani Khalili
Ali Nemati
and Ali Ghorbani
Departemant of Veterinary, University of Shahed Chamran Ahwaz, Khuzestan
Departemant of Animal Science, Islamic Azad University Khorasgan, Isfahan
Departemant of Poultry Nutrition, Islamic Azad University Karaj, Alborz
Departemant of Animal Science, Islamic Azad University Shahrekord, Chaharmahal Bakhtiary
Department of Animal Science, Islamic Azad UniversityScience and Research Branch, Tehran, Iran
Contamination of food and livestock feed to common poisonous fungus in the air and environment is important.
Contaminated animal feed to a atoxin producing fungus leads to the disturbance in the cycle of animal health, milk
and consumers of animal products. In this study, all isolates of Aspergillus on food have been studied with the aim
of identifying and toxin-producing. Aspergillus are a group of fungus that have replication power and abundant
growth, and are known as a part of fungal  ora of most of places, and among Aspergillus, a number of species have
capability of toxin-producing and can create disease in humans and animals. In this study, 180 samples of poultry in
the city of Ahvaz were prepared and after transfer to the laboratory and culture, puri cation was performed, then the
identi cation of isolates using the valid keys of mycology was done and extracting toxin of species on the context
of corn was also performed, also determining toxin-producing of species was done in two ways of coconut agar and
TLC. The results showed in this study several species of Aspergillus, including species of flavus A. A.parasitichus,
A.ochraceus, A.niger were identi ed. Species avus with 52 isolates (0.028) allocated the most frequency and species
A.ochraceus with 9 isolates (4.98) had the lowest frequency. Informing toxin-producing of the fungus on poultry diet
and how toxin-producing of this fungus can be helpful in the management and prevention of infection by the fungus.
*Corresponding Author:
Received 27
Dec, 2016
Accepted after revision 2
March, 2017
BBRC Print ISSN: 0974-6455
Online ISSN: 2321-4007
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Biosci. Biotech. Res. Comm. Special Issue No 1:272-275 (2017)
Abdoallah Beatsayah et al.
Fungus are present in abundant in the air, soil and
our environment, so in the presence of humidity and
the temperature, growth and reproduction of fungi are
escalated (Ersali et al., 2008). Some of the toxins in the
environment are fungal toxins that a atoxins are con-
sidered the most important of them (Aghababaei et al.,
2012). A atoxins are among the most important myco-
toxins produced by some Aspergillus species (Aspergil-
lus  avus, Aspergillus parasiticus) and Penicillium (Pen-
icillium Pvbrvlvm) (Jamali Emam Ghedis and Moeini.,
Aspergillus is fungal infectious disease of young
poultry and chronic disease of older poultry that occurs
with breathing dif culties. Fumigatus is the cause of
Aspergillus fungus that remains in the nature by pro-
ducing spore in nature. Fungus grows and reproduces
rapidly in temperature 37 degrees and proper humidity
but the increase of humidity causes the stop of growth.
Considering its growth speed, it eliminates one-day
chicks earlier that have lower resistance. Disease course
varies depending on the age and power of poultry that
in young poultry in acute form takes one week and in
old poultry in chronic form takes a few weeks. The exist-
ence of some fungi in feed for livestock and poultry is
natural and if they not have uncontrolled growth, they
not have health risks but some fungi that are called fun-
gus of toxin-producing have particular importance.
Mycotoxins are secondary metabolites produced by
some  lamentous fungi that infect agricultural products.
They are toxic for humans and animals and causing a
signi cant reduction in product returns and economic
losses. There are widespread reports of outbreak of fun-
gal damages in different countries. The metabolites are
produced by different species under special conditions in
terms of temperature, humidity and oxygen and are not
essential for cellular activities. Consumption of foods
contaminated with mycotoxins has been associated with
a number of human poisoning, and even poisoning with
mycotoxins can sometimes cause death (Aghababaei
et al., 2012).
Peanuts, pistachios, wheat, rice, corn, almonds and
gs are the main hosts of the fungus and are known
as the most perfect natural environment for the growth
of a atoxin-producing fungi in the world, yet, several
types of a atoxin have been detected that among them
B1, B2, G1, G have the utmost importance in the world.
Among the different types of a atoxin known, a atoxin
B1 by the International Agency for carcinogens is placed
in Group A, research on cancer and in the meantime the
toxicity and carcinogenicity of a atoxin B1 is reported
over other types, contamination of feed for livestock and
poultry to fungus, followed by it a atoxins, in addition
to economic losses jeopardizes consumers of food with
animal origin. Therefore, tracking and evaluation of
a atoxins in food and animal feed and comparison it
with standard values in order to knowledge, proposals
and measures to prevent A atoxicosis in animals and
humans is necessary. Several studies are conducted on
investigating the presence of a atoxins in feed in Iran
and many countries (Kan and Meijer, 2007).
Clinical signs of a atoxin toxicity include: autopsy
lesions, autopsy injuries, histopathological lesions in
tissue, as well as the effects created on production indi-
ces of poultry  ocks in cases of experimental and nat-
ural occurrence of A atoxicosis in broiler chickens in
worldwide is reported (Kanungo et al., 2011). The main
body attacked by the a atoxin is liver and in human
causes severe disorders of the liver occur. In animals
also it causes problems in the gut, preventing immune
function, reducing reproduction, increasing feed con-
version ef ciency, reducing the production of milk and
egg, anemia, jaundice, reducing growth. Considering the
importance of toxin-producing spices on food and diets
of poultry in order to recognize species of toxin-pro-
ducing and how their toxin-producing seems necessary.
In order to identify isolates of Aspergillus fungus in
poultry in Ahvaz during 2015-2016, the sampling was
done in 4 active poultry farms in different parts of the
city of Ahvaz. Samples taken were formed of various
materials such as all diets used by poultry including:
starter, middle-feeding and post-feeding, corn and soy-
beans, 150 samples were separated and placed inside
plastic bags in the refrigerator at a temperature 4 ° C.
Then the potato dextrose agar medium for culturing and
isolation of fungi from sample was used. Samples were
placed for 24 to 48 hours in incubation at 25 ° C then for
purifying fungus, new colonies grown were cultivated.
A. Puri cation of fungus
In order to purify a fungal isolates, single spore method
was used. So that by Anas  ne needle, a small amount
of Fungi spore was transferred to tube containing
distilled water and then by Lam Thomas, its dilution was
calculated. After preparing a suitable dilution, diameters
of the suspension was transferred to medium of water
agar 2% and by passing 12 to 24 hours, the colonies
grown of fungus on medium (PDA) that already poured
and cooled in the three spots was transferred and for 5
to 7 days at 25 ° C were reared (
Lanyasunya et al., 2005).
B. Identi cation of fungus
To identify fungus, different culture media was used,
including the culture of potato, dextrose, agar (PDA) is
Abdoallah Beatsayah et al.
Table 1. The sample and the number of food rations contaminated with Aspergillus fungus
number and percent of sample
A Flavus.A. Parasitichu.A. Niger.A. ochraceus
Percent of contamination300.0280.01822.254.98
known a public culture medium and used for the cul-
tivation and primary isolation of isolates, as well as to
identify the isolates, speci c culture mediums such as
CY20S (Czapek Yeast Extract Agar with20 sucrose) were
C. Preparing context of corn
Value of 100 grams of corn separately was poured in
separate Erlenmeyer and the ratio of 100 to 27 was
added to each of the contexts of distilled water and in
temperature of 121 ° C. and 1.5 atmospheric pressure for
15 minutes was sterilized in an autoclave and 24 hours
later was autoclaved again to all the external factors to
be eliminated.
C. Inoculum of fungus to contexts of culture
After puri cation fungus, the fungus was developed on
PDA medium and then spore suspension was formed in
distilled water and 10 cc of this suspension in sterile
conditions under the hood of Laminar was added to
context and  asks with treatments were placed at room
temperature and the conditions 24 hours of light and
D. Extraction of toxin
In order to extract Mycotoxins,  rst the contents of the
asks for 24 hours at 73 ° C were placed and then milled
samples contaminated seeds acetonitrile - methanol
was added and then was placed for 3 hours on a shaker.
The resulting mixture was smooth with Whatman  lter
paper and extract was collected in clean Falcons and
extract was passed from puri cation column containing
a cationic resin and alumina-Carbon mix. For this
purpose, the beginning and the end of the column with
a layer of glass wool is blocked and a gram of mixture
of Almunia - carbon is added on it and again a layer
of glass wool blocked is placed on it and then cation
resin that already for 1 hour is placed in distilled water
is poured in column and then extract of samples were
passed through the column.
Passed extract of  rst column using the second puri-
cation column includes respectively glass wool and
alumina-carbon and net glass wool and then extract
passed through the second puri cation column for sol-
vent evaporation was transferred to oven 70 °. After
drying the extract, methanol-water solution is added to
it and for an hour is placed at room temperature, then
methanol-water and acetonitrile-methanol solution was
added and in order to evaporate, the solvent again is
placed in an oven at 73 ° and  nally methanol-water
was added to dried extract and the extracts were stored
at -20 ° C (Rahimi et al., 2011).
E. Investigating toxin-producing by T LC
After extracting the toxins, solutions of sample and
a atoxin standard on TLC of TLC aluminum plate with
silica gel in a direction were dotted, after the plate was
placed in TLC tank and methanol: acetonitrile (88:12)
risen and the plate is washed up. Plate is dried in the
air and under the lamp UV (365 nm) will be judged and
blue  uorescence intensity is compared with standard
uorescence (
Rahimi et al., 2008).
The results of contamination of food rations including
corn, soybean, starter, middle-feeding, post-feeding and
context of poultry to Aspergillus fungus are in Table
1. The results showed in this study several species of
Aspergillus, including species A.  avus A. parasitichus,
A.ochraceus, A.niger were identi ed. Species of avus
with 52 isolates (0.028) allocated the most frequency and
the species A.ochraceus with 9 isolates (4.98) had the
lowest frequency. The presence of fungi such as A. avus
with the most frequency among Aspergillus species can
be important in terms of toxin-producing; this fungus
can produce a atoxin B1 that has the highest toxicity
than other A atoxin produced by Aspergillus. As well
as there are species of other toxin-producing on diet of
poultry that including them can be noted to Aspergillus
Abdoallah Beatsayah et al.
parasiticus fungi. The species has the ability to produce
each 4 kinds of a atoxin B1, B2, G1, G2, and in this
sense can be considered a dangerous toxin-producing
Ochratoxin A is one of mycotoxins produced by the
fungi Aspergillus and Penicillium. Spore of this fungus
are widely dispersed in the environment and a lot of food,
especially cereals are infected these fungus (Thompson
and Henke. 2000) hence the toxin-producing spices in
poultry as a pathogen agent could jeopardize poultry
health. Since the existence of the toxin-producing spices
on the dietary food for poultry and food can be danger-
ous, so the availability of toxin-producing agents and
how toxin-producing is very important, Aspergillus is
the cause of A atoxicosis in poultry as well as carcino-
genic in human (
Kanungo et al., 2011).
Brugfer et al (2003) in Australia on wheat and wheat
our used in animal feed of farms studied isolated and
identi ed molds of Aspergillus, Cladosporydyom and
Penicillium, the type of fungi identi ed in this study
con rms the results of previous studies. Among the feed
of livestock, corn silage and concentrate, respectively,
had the highest number of fungal colonies. In a study
that was done in Mazandaran, corn has the highest
percentage to fungus Aspergillus  avus (
Rahimi et al.,
2008). This study also has identi ed corn silage as a
feed contaminated with Aspergillus  avus that hence,
con rms previous studies, but in this study, the most
frequency of this type of fungus has been observed in
straw and this could be due to difference in preparation
and storage conditions of animal feeds in different farms
and cities.
In our country, according to the climatic variation, kinds
of toxin-producing strains of aspergillus can contami-
nate food and cause huge physical and  nancial losses.
Also, due to diverse conditions of weather, there is the
possibility of a wide range of fungi pruducing myco-
toxins with toxins relevant in the environment, and it
should be done sanitation and prevention principles as
well as compliance with international standards for the
maintenance and storage of goods and food rations.
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